Name: induction and diagnosis of tumors in drosophila imaginal disc epithelia
Uploaded: 2017-07-25T14:00:00
Description: Watch this Scientific Journal Video about Induction and Diagnosis of Tumors in Drosophila Imaginal Disc Epithelia at JoVE.com

我们感谢J. Vaughen对手稿的批判性阅读。这项工作得到了JSPS KAKENHI Grant号26891025,15H01500和武田科学基金会YT研究奖学金的资助

飞越和克隆诱导<ol><li>在收集原始苍蝇之前12小时内取出小瓶中的所有苍蝇。 </li><li>通过注入CO 2气体和地方飞行到CO 2蝇垫麻醉在小瓶中的苍蝇。 </li><li>将10-20名女性和10名男性从CO 2飞虱转移到新鲜的小瓶中，并在25℃下孵育1天。 </li><li>将这些苍蝇转移到新鲜的小瓶中，并在25℃下孵育12小时。 注意：丢弃第一个小瓶，因为女性在第一天没有铺好足够的鸡蛋。 </li><li>...

<ol>
	<li>Hanahan, D., Weinberg, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+hallmarks+of+cancer.">The hallmarks of cancer.</a> Cell. 100 (1), 57-70 (2000).</li><li>Xu, T., Rubin, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+genetic+mosaics+in+developing+and+adult+Drosophila+tissues.">Analysis of genetic mosaics in developing and adult Drosophila tissues.</a> Development. 117 (4), 1223-1237 (1993).</li><li>Struhl, G., Basler, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Organizing+activity+of+wingless+protein+in+Drosophila.">Organizing activity of wingless protein in Drosophila.</a> Cell. 72 (4), 527-540 (1993).</li><li>Potter, C. J., Turenchalk, G. S., Xu, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+in+cancer+research.+An+expanding+role.">Drosophila in cancer research. An expanding role.</a> Trends Genet. 16 (1), 33-39 (2000).</li><li>Miles, W. O., Dyson, N. J., Walker, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modeling+tumor+invasion+and+metastasis+in+Drosophila.">Modeling tumor invasion and metastasis in Drosophila.</a> Dis Model Mech. 4 (6), 753-761 (2011).</li><li>Tipping, M., Perrimon, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+as+a+model+for+context-dependent+tumorigenesis.">Drosophila as a model for context-dependent tumorigenesis.</a> J Cell Physiol. 229 (1), 27-33 (2013).</li><li>Bilder, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+polarity+and+proliferation+control:+links+from+the+Drosophila+neoplastic+tumor+suppressors.">Epithelial polarity and proliferation control: links from the Drosophila neoplastic tumor suppressors.</a> Genes Dev. 18 (16), 1909-1925 (2004).</li><li>Humbert, P. O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Control+of+tumourigenesis+by+the+Scribble/Dlg/Lgl+polarity+module.">Control of tumourigenesis by the Scribble/Dlg/Lgl polarity module.</a> Oncogene. 27 (55), 6888-6907 (2008).</li><li>Huang, L., Muthuswamy, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polarity+protein+alterations+in+carcinoma:+a+focus+on+emerging+roles+for+polarity+regulators.">Polarity protein alterations in carcinoma: a focus on emerging roles for polarity regulators.</a> Curr Opin Genet Dev. 20 (1), 41-50 (2010).</li><li>Brumby, A. M., Richardson, H. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=scribble+mutants+cooperate+with+oncogenic+Ras+or+Notch+to+cause+neoplastic+overgrowth+in+Drosophila.">scribble mutants cooperate with oncogenic Ras or Notch to cause neoplastic overgrowth in Drosophila.</a> EMBO J. 22 (21), 5769-5779 (2003).</li><li>Igaki, T., Pastor-Pareja, J. C., Aonuma, H., Miura, M., Xu, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intrinsic+tumor+suppression+and+epithelial+maintenance+by+endocytic+activation+of+Eiger/TNF+signaling+in+Drosophila.">Intrinsic tumor suppression and epithelial maintenance by endocytic activation of Eiger/TNF signaling in Drosophila.</a> Dev Cell. 16 (3), 458-465 (2009).</li><li>Tamori, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+Lgl+and+Mahjong/VprBP+in+cell+competition.">Involvement of Lgl and Mahjong/VprBP in cell competition.</a> PLoS Biol. 8 (7), e1000422 (2010).</li><li>Cordero, J. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oncogenic+Ras+diverts+a+host+TNF+tumor+suppressor+activity+into+tumor+promoter.">Oncogenic Ras diverts a host TNF tumor suppressor activity into tumor promoter.</a> Dev Cell. 18 (6), 999-1011 (2010).</li><li>Yamamoto, M., Ohsawa, S., Kunimasa, K., Igaki, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+ligand+Sas+and+its+receptor+PTP10D+drive+tumour-suppressive+cell+competition.">The ligand Sas and its receptor PTP10D drive tumour-suppressive cell competition.</a> Nature. 542 (7640), 246-250 (2017).</li><li>Vaughen, J., Igaki, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Slit-Robo+Repulsive+Signaling+Extrudes+Tumorigenic+Cells+from+Epithelia.">Slit-Robo Repulsive Signaling Extrudes Tumorigenic Cells from Epithelia.</a> Dev Cell. 39 (6), 683-695 (2016).</li><li>Tamori, Y., Suzuki, E., Deng, W. -. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+Tumors+Originate+in+Tumor+Hotspots,+a+Tissue-Intrinsic+Microenvironment.">Epithelial Tumors Originate in Tumor Hotspots, a Tissue-Intrinsic Microenvironment.</a> PLoS Biol. 14 (9), e1002537 (2016).</li><li>Ohsawa, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elimination+of+oncogenic+neighbors+by+JNK-mediated+engulfment+in+Drosophila.">Elimination of oncogenic neighbors by JNK-mediated engulfment in Drosophila.</a> Dev Cell. 20 (3), 315-328 (2011).</li><li>Men&#233;ndez, J., P&#233;rez-Garijo, A., Calleja, M., Morata, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+tumor-suppressing+mechanism+in+Drosophila+involving+cell+competition+and+the+Hippo+pathway.">A tumor-suppressing mechanism in Drosophila involving cell competition and the Hippo pathway.</a> Proc Natl Acad Sci U S A. 107 (33), 14651-14656 (2010).</li><li>Hogan, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+the+interface+between+normal+and+transformed+epithelial+cells.">Characterization of the interface between normal and transformed epithelial cells.</a> Nat Cell Biol. 11 (4), 460-467 (2009).</li><li>Kajita, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interaction+with+surrounding+normal+epithelial+cells+influences+signalling+pathways+and+behaviour+of+Src-transformed+cells.">Interaction with surrounding normal epithelial cells influences signalling pathways and behaviour of Src-transformed cells.</a> J Cell Sci. 123 (Pt 2), 171-180 (2010).</li><li>Norman, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Loss+of+Scribble+causes+cell+competition+in+mammalian+cells.">Loss of Scribble causes cell competition in mammalian cells.</a> J Cell Sci. 125 (1), 59-66 (2012).</li><li>Wagstaff, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+cell+competition+kills+cells+via+induction+of+lethal+p53+levels.">Mechanical cell competition kills cells via induction of lethal p53 levels.</a> Nat Commun. 7, 1-14 (2016).</li><li>Kajita, M., Fujita, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=EDAC:+Epithelial+defence+against+cancer-cell+competition+between+normal+and+transformed+epithelial+cells+in+mammals.">EDAC: Epithelial defence against cancer-cell competition between normal and transformed epithelial cells in mammals.</a> J Biochem. 158 (1), 15-23 (2015).</li><li>North, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Seeing+is+believing?+A+beginners'+guide+to+practical+pitfalls+in+image+acquisition.">Seeing is believing? A beginners' guide to practical pitfalls in image acquisition.</a> J Cell Biol. 172 (1), 9-18 (2006).</li><li>Schindelin, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fiji:+an+open-source+platform+for+biological-image+analysis.">Fiji: an open-source platform for biological-image analysis.</a> Nat Methods. 9 (7), 676-682 (2012).</li><li>Brand, A. H., Perrimon, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+gene+expression+as+a+means+of+altering+cell+fates+and+generating+dominant+phenotypes.">Targeted gene expression as a means of altering cell fates and generating dominant phenotypes.</a> Development. 118 (2), 401-415 (1993).</li><li>Jory, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+survey+of+6,300+genomic+fragments+for+cis-regulatory+activity+in+the+imaginal+discs+of+Drosophila+melanogaster.">A survey of 6,300 genomic fragments for cis-regulatory activity in the imaginal discs of Drosophila melanogaster.</a> Cell Rep. 2 (4), 1014-1024 (2012).</li><li>McGuire, S. E., Le, P. T., Osborn, A. J., Matsumoto, K., Davis, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatiotemporal+rescue+of+memory+dysfunction+in+Drosophila.">Spatiotemporal rescue of memory dysfunction in Drosophila.</a> Science. 302 (5651), 1765-1768 (2003).</li><li>Rodrigues, A. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activated+STAT+regulates+growth+and+induces+competitive+interactions+independently+of+Myc,+Yorkie,+Wingless+and+ribosome+biogenesis.">Activated STAT regulates growth and induces competitive interactions independently of Myc, Yorkie, Wingless and ribosome biogenesis.</a> Development. 139 (21), 4051-4061 (2012).</li><li>Khan, S. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+neoplasia+in+Drosophila+entails+switch+to+primitive+cell+states.">Epithelial neoplasia in Drosophila entails switch to primitive cell states.</a> Proc Natl Acad Sci U S A. 110 (24), E2163-E2172 (2013).</li><li>Pagliarini, R. A., Xu, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+genetic+screen+in+Drosophila+for+metastatic+behavior.">A genetic screen in Drosophila for metastatic behavior.</a> Science. 302 (5648), 1227-1231 (2003).</li><li>Gonzalez, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+melanogaster:+a+model+and+a+tool+to+investigate+malignancy+and+identify+new+therapeutics.">Drosophila melanogaster: a model and a tool to investigate malignancy and identify new therapeutics.</a> Nat Rev Cancer. 13 (3), 172-183 (2013).</li><li>Patel, P. H., Edgar, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue+design:+how+Drosophila+tumors+remodel+their+neighborhood.">Tissue design: how Drosophila tumors remodel their neighborhood.</a> Semin Cell Dev Biol. 28, 86-95 (2014).</li><li>Nakajima, Y. -. I., Meyer, E. J., Kroesen, A., McKinney, S. A., Gibson, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+junctions+maintain+tissue+architecture+by+directing+planar+spindle+orientation.">Epithelial junctions maintain tissue architecture by directing planar spindle orientation.</a> Nature. 500 (7462), 359-362 (2013).</li><li>Colombani, J., Andersen, D. S., L&#233;opold, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Secreted+peptide+Dilp8+coordinates+Drosophila+tissue+growth+with+developmental+timing.">Secreted peptide Dilp8 coordinates Drosophila tissue growth with developmental timing.</a> Science. 336 (6081), 582-585 (2012).</li><li>Garelli, A., Gontijo, A. M., Miguela, V., Caparros, E., Dominguez, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaginal+discs+secrete+insulin-like+peptide+8+to+mediate+plasticity+of+growth+and+maturation.">Imaginal discs secrete insulin-like peptide 8 to mediate plasticity of growth and maturation.</a> Science. 336 (6081), 579-582 (2012).</li></ol>

的GAL4-UAS系统是用于在果蝇 26靶基因表达的最有力的遗传工具之一，极大地方便了肿瘤细胞的诱导和分析在体内 4。该系统能够产生携带敲低肿瘤抑制基因的克隆或在野生型上皮组织内癌基因的过度表达，这种情况与人类癌症的初始阶段非常相似，其中转化的前肿瘤细胞被正常上皮细胞包围。由GAL4增强子捕获系或Janelia GAL4系27等增...

上皮组织是高度组织的系统，具有显着的稳态能力，通过发育和细胞周转维持其组织。然而，这种健壮的自组织系统在肿瘤发育过程中逐渐被破坏。在肿瘤发展开始时，由癌基因激活或肿瘤抑制基因失活引起的单个突变细胞出现在上皮层内。当这种转化的“前肿瘤细胞”逃避抑制环境时，会破坏上皮组织，并开始不受控制的增殖，发生肿瘤发生1 。在过去几十年中，遗传学和分子生物学方面的杰出技术进步在癌症研究方面取得了显着进展。特别是最近在黑腹果蝇中使用遗传马赛克分析工具的研究 ，如FLP-FRT（flippase重组酶/翻转酶重组酶靶）有丝分裂重组通货膨胀2和翻转出-GAL4-UAS（上游激活序列）系统3，已经极大地促进了更好地理解参与了肿瘤4，5，6的形成和转移的遗传机制。 一组保守的果蝇肿瘤抑制基因， 致死巨幼虫 （ lgl ）， 圆盘大 （ dlg ）和涂鸦 （ scrib ）的研究突出了上皮组织损失和肿瘤发展之间的关键关系，因为这些基因发挥关键作用在上皮组织中调节顶端细胞极性和细胞增殖7 。虽然果蝇成像圆盘通常是单层上皮，这三个基因中的任何一个的纯合突变导致细胞失去结构和极性，不能形成差异过度膨胀，最终形成与相邻组织融合的多层无定形块7 。同样，这些基因在哺乳动物的破坏是参与恶性肿瘤8,9的发展。由突变体组织表现出的表型的肿瘤已导致这三个基因的分类为保守的，肿瘤性的肿瘤抑制基因（nTSGs）7,8。然而，当纯合nTSG突变细胞开发使用FLP-FRT-介导的有丝分裂重组的野生型成虫盘被零散地生成，突变的细胞从组织至c-Jun N-末端激酶（JNK） -依赖性的细胞凋亡10,11消除，12，13，14，15挤压</suP&gt;， 如图16所示 ，或通过邻居17吞没和吞噬作用。在该镶嵌基因的上皮细胞凋亡主要是在位于克隆边界nTSG突变体细胞中检测到，这表明相邻正常细胞触发nTSG突变的细胞10，11，12，18的细胞凋亡。在哺乳动物细胞中最近的研究已经证实，有利于肿瘤细胞的这种细胞依赖性竞争淘汰是对抗癌症的19，20，21，22，23的进化上保守的上皮细胞的自我防御机制。 然而，最近对果蝇成像圆盘的研究表明，镶嵌nTSG敲低克隆诱导特异性的肿瘤性肿瘤机翼成像光盘的ic区域16 。初始肿瘤形成始终存在于外周“铰链”区域，并且从未在翼盘上皮的中央“囊”区域观察到，这表明nTSG敲低细胞的致瘤潜力取决于局部环境。中央袋区域作为“肿瘤冷点”，其中肿瘤细胞不显示发育不良的过度生长，而外周铰链区则表现为“肿瘤热点” 16 。在“冷点”小袋区域，nTSG敲低细胞从基底侧分层并进行细胞凋亡。相比之下，由于“热点”铰链细胞在其基底具有强大的细胞骨架结构网络，所以nTSG敲低细胞从上皮的顶端分层并引发致瘤性过度生长16 。因此，成像光盘中肿瘤表型的分析需要仔细考虑区域特异性致瘤性刺激易感性的降容。 在这里，我们描述了使用GAL4-UAS- RNAi系统在果蝇翼成像盘中诱导肿瘤肿瘤形成的方案，通过其在正常翼盘上皮细胞中产生nTSG敲低细胞。虽然这些实验系统对于研究癌症的早期阶段是有用的，但是尚未清楚地描述用于评估成像盘上皮细胞肿瘤进展阶段的明确分类方法。因此，我们还提出了一种将翼片上皮诱导的前肿瘤克隆表型分为三类：增生（增殖增多的正常出现细胞数量过多），异常增生（异常出现的恶化前组织细胞）和瘤形成（良性或恶性肿瘤由具有异常外观和异常增殖模式的细胞组成）。

<table>
	<tbody>
		<tr>
			<td>Reagents</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate buffered saline (PBS)</td>
			<td>Wako</td>
			<td>162-19321</td>
			<td></td>
		</tr>
		<tr>
			<td>TritonX-100</td>
			<td>Wako</td>
			<td>168-11805</td>
			<td></td>
		</tr>
		<tr>
			<td>Formaldehyde</td>
			<td>Wako</td>
			<td>064-00406</td>
			<td></td>
		</tr>
		<tr>
			<td>bovine serum albumin</td>
			<td>Sigma</td>
			<td>A7906</td>
			<td></td>
		</tr>
		<tr>
			<td>normal goat serum</td>
			<td>Sigma</td>
			<td>G6767</td>
			<td></td>
		</tr>
		<tr>
			<td>mounting medium, Vectashield</td>
			<td>Vector Laboratories</td>
			<td>H-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>DAPI</td>
			<td>Sigma</td>
			<td>D9542</td>
			<td></td>
		</tr>
		<tr>
			<td>mouse-anti-Dlg 4F3</td>
			<td>Developmental Studies Hybridoma Bank</td>
			<td>4F3 anti-discs large, RRID:AB_528203</td>
			<td>dilute in PBTG, 1:40</td>
		</tr>
		<tr>
			<td>mouse-anti-MMP1</td>
			<td>Developmental Studies Hybridoma Bank</td>
			<td>3A6B4, RRID:AB_579780</td>
			<td>3 mixed 1:1:1 and dilute in PBTG, 1:40</td>
		</tr>
		<tr>
			<td>mouse-anti-MMP1</td>
			<td>Developmental Studies Hybridoma Bank</td>
			<td>3B8D12, RRID:AB_579781</td>
			<td>3 mixed 1:1:1 and dilute in PBTG, 1:40</td>
		</tr>
		<tr>
			<td>mouse-anti-MMP1</td>
			<td>Developmental Studies Hybridoma Bank</td>
			<td>5H7B11, RRID:AB_579779</td>
			<td>3 mixed 1:1:1 and dilute in PBTG, 1:40</td>
		</tr>
		<tr>
			<td>mouse-anti-atubulin</td>
			<td>Developmental Studies Hybridoma Bank</td>
			<td>AA4.3, RRID:AB_579793</td>
			<td>dilute in PBTG, 1:100</td>
		</tr>
		<tr>
			<td>Alexa Fluor 546 Phalloidin</td>
			<td>Molecular probes</td>
			<td>A22283</td>
			<td>dilute in PBS, 1:40</td>
		</tr>
		<tr>
			<td>goat anti-mouse IgG antibody, Alexa Fluor 546</td>
			<td>Molecular probes</td>
			<td>A11030</td>
			<td>dilute in PBTG, 1:400</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Fly strains</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>sd-Gal4</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#8609</td>
			<td>recombined with UAS-EGFP</td>
		</tr>
		<tr>
			<td>upd-Gal4</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#26796</td>
			<td>recombined with UAS-EGFP</td>
		</tr>
		<tr>
			<td>UAS-lgl-RNAi</td>
			<td>Vienna Drosophila RNAi center</td>
			<td>#51247</td>
			<td></td>
		</tr>
		<tr>
			<td>UAS-scrib-RNAi</td>
			<td>Vienna Drosophila RNAi center</td>
			<td>#105412</td>
			<td></td>
		</tr>
		<tr>
			<td>UAS-RasV12</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#64196</td>
			<td></td>
		</tr>
		<tr>
			<td>UAS-Yki3SA</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#28817</td>
			<td></td>
		</tr>
		<tr>
			<td>hsFLP</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#6</td>
			<td></td>
		</tr>
		<tr>
			<td>Act&#62;CD2&#62;GAL4 (flip-out GAL4)</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#4780</td>
			<td>recombined with UAS-EGFP</td>
		</tr>
		<tr>
			<td>UAS-EGFP</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#5428</td>
			<td>X chromosome</td>
		</tr>
		<tr>
			<td>UAS-EGFP</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#6658</td>
			<td>third chromosome</td>
		</tr>
		<tr>
			<td>UAS-Dicer2</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#24650</td>
			<td>second chromosome</td>
		</tr>
		<tr>
			<td>UAS-Dicer2</td>
			<td>Bloomington Drosophila Stock Center</td>
			<td>#24651</td>
			<td>third chromosome</td>
		</tr>
		<tr>
			<td>vkg-GFP</td>
			<td>Morin et al. 2001</td>
			<td></td>
			<td>GFP protein trap</td>
		</tr>
	</tbody>
</table>

induction and diagnosis of tumors in drosophila imaginal disc epithelia

在癌症的早期阶段，转化的突变细胞显示细胞学异常，开始不受控制的过度生长，并逐渐破坏组织组织。 黑腹果蝇已成为癌症生物学中流行的实验模型系统，用于研究肿瘤发生的遗传和细胞机制。特别地，用于果蝇成像盘（在幼虫中发育的上皮）的遗传工具使得能够在正常上皮组织内形成转化的前肿瘤细胞，这种情况类似于人类癌症的初始阶段。然而，最近关于果蝇翼成像圆盘中的肿瘤发生的研究表明，肿瘤起始取决于组织固有的细胞结构和局部微环境，这表明在评估肿瘤表型的成像中考虑到致瘤性刺激的区域特异性易感性是重要的光盘。促进肿瘤进展的表型分析在成像光盘中，我们描述了使用GAL4-UAS系统在机翼成像光盘中诱导肿瘤性肿瘤的遗传实验方案。我们进一步介绍一种诊断方法来分类在上皮细胞中诱导的克隆性病变的表型，因为以前没有描述过用于区分肿瘤进展的各个阶段（如增生，发育不良或瘤形成）的明确分类方法。这些方法可能广泛适用于果蝇各种器官的肿瘤表型的克隆分析。

为了证实通过RNAi介导的果蝇翼成像盘中的RNAi介导的nTSG敲低实验诱导的肿瘤肿瘤形成，使用三种不同的GAL4驱动子来表达用于lgl或scrib的 UAS-RNAi ：（1） sd-GAL4，其驱动强的UAS表达翼袋和铰链区域的轻度表达（ 图2和图3A ）; （2）在铰链子域中驱动的upd-GAL4，包括在背内?...

Watch this Scientific Journal Video about Induction and Diagnosis of Tumors in Drosophila Imaginal Disc Epithelia at JoVE.com

Induction and Diagnosis of Tumors in Drosophila Imaginal Disc Epithelia

果蝇成像盘上皮的马赛克克隆分析是研究肿瘤发生的遗传和细胞机制的有力模型系统。在这里，我们描述了使用GAL4-UAS系统在果蝇翼成像盘中诱导肿瘤的方案，并介绍了分类肿瘤表型的诊断方法。

诱导和诊断肿瘤<em&gt;果蝇</em&gt; Imaginal Disc Epithelia

In the early stages of cancer, transformed mutant cells show cytological abnormalities, begin uncontrolled overgrowth, and progressively disrupt tissue ...

Research

JoVE Journal

Cancer Research

Induction and Diagnosis of Tumors in Drosophila Imaginal Disc Epithelia

Next Generation Sequencing for the Detection of Actionable Mutations in Solid and Liquid Tumors

新一代测序为可操作的突变在固体和液体瘤检测

As our understanding of the driver mutations necessary for initiation and progression of cancers improves, we gain critical information on how specific ...

科研

癌症研究

The Drosophila Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics

The Drosophila Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics

该<em&gt;果蝇</em&gt;成虫盘瘤模型：基因的表达与肿瘤侵袭性的可视化和定量遗传马赛克

Drosophila melanogaster has emerged as a powerful experimental system for functional and mechanistic studies of tumor development and progression in the ...

Development of a Human Preclinical Model of Osteoclastogenesis from Peripheral Blood Monocytes Co-cultured with Breast Cancer Cell Lines

乳腺癌细胞系外周血单核细胞破的人前临床模型的建立

The crosstalk between tumor cells and bone cells in the bone microenvironment is crucial to understanding the mechanism of bone metastasis formation. We ...

Establishment of a Primary Culture of Patient-derived Soft Tissue Sarcoma

病人衍生软组织肉瘤原代培养的建立

Soft tissue sarcomas (STS) represent a spectrum of heterogeneous malignancies with a difficult diagnosis, classification, and management. To date, more ...

Enrichment and Characterization of the Tumor Immune and Non-immune Microenvironments in Established Subcutaneous Murine Tumors

肿瘤免疫和非免疫微环境在小鼠皮下肿瘤中的富集与鉴定

The tumor immune microenvironment (TIME) has recently been recognized as a critical mediator of treatment response in solid tumors, especially for ...

Murine Model of Metastatic Liver Tumors in the Setting of Ischemia Reperfusion Injury

缺血再灌注损伤中转移性肝肿瘤的模样

Liver ischemia and reperfusion (I/R) injury, a common clinical challenge, remains an inevitable pathophysiological process that has been shown to induce ...

Obtaining Cancer Stem Cell Spheres from Gynecological and Breast Cancer Tumors

从妇科和乳腺癌肿瘤获得癌症干细胞球

Cancer stem cells (CSC) are a small population with self-renewal and plasticity which are responsible for tumorigenesis, resistance to treatment and ...

Orthotopic Transplantation of Breast Tumors as Preclinical Models for Breast Cancer

乳腺癌临床前模型乳腺肿瘤的矫形移植

Preclinical models that faithfully recapitulate tumor heterogeneity and therapeutic response are critical for translational breast cancer research. ...

Quail Chorioallantoic Membrane - A Tool for Photodynamic Diagnosis and Therapy

The chorioallantoic membrane (CAM) of an avian embryo is a thin, extraembryonic membrane that functions as a primary respiratory organ. Its properties ...

High-Resolution Ultrasonography for the Analysis of Orthotopic ATC Tumors in a Genetically Engineered Mouse Model

用于分析基因工程小鼠模型中原位ATC肿瘤的高分辨率超声检查

Anaplastic thyroid carcinoma (ATC) is associated with a poor prognosis and short median survival time, but no effective treatment improves the outcomes ...